PROJECT SUMMARY Branched tissue networks are conserved throughout the animal kingdom, often found in organs that require a large surface area relative to a confining tissue volume. The lung uses one such branched epithelial network to achieve rapid gas exchange. Deviation from the normal branching morphogenesis of the lung airways results in respiratory defects, such as pulmonary hypoplasia. Normal airway epithelial morphogenesis requires three critical factors, including branch site specification, branch elongation, and mesenchymal remodeling to shape the elongating branch. Therefore, understanding mechanisms that influence branching patterns will provide a molecular and physical basis for bronchopulmonary abnormalities. The role of the pulmonary mesenchyme in directing branch formation by secreting diffusible factors, such as fibroblast growth factors (FGFs), is well established. However, how mesenchymal cells alter the mechanical properties of the microenvironment adjacent to the airway epithelium is less clear. Epithelial branching in the mammary and salivary glands requires the turnover of basement membrane (BM) proteins at branch tips to promote elongation, presumably because intact BM constrains epithelial growth. BM proteins appear to be reduced at the tips of airway epithelial branches, although it is not clear how constraints imposed by the BM may affect airway branch initiation. In the mammalian lung, airway smooth muscle (SM) differentiates at sites of epithelial branch bifurcation and contractions of SM are required for branching morphogenesis. While SM contractions are required for normal airway branching, it is unclear how SM differentiation is specified at branch tips. This proposal aims to illuminate how the extracellular matrix (ECM) and SM contractions constrain epithelial branches to direct branching morphogenesis. This work will provide insight into how mesenchymal remodeling may alter the mechanical microenvironment proximal to epithelial branches in the lung, and thereby direct airway branching morphogenesis.